Construction and characterization of a recombinant fowlpox virus containing HIV-1 multi-epitope-p24 chimeric gene in mice

Abstract

The epidemic of HIV/AIDS is sweeping across the world. It is of great importance to figure out new ways to curb this disease. Epitope-based vaccine is one of these solutions. In this study, a chimeric gene was obtained by combination of a designed HIV-1 multi-epitope gene (MEG) and HIV-1 p24 gene. A recombinant plasmid pUTA2-MEGp24 was then constructed by inserting MEGp24 gene into the downstream of the promoter (ATI-P7.5x20) of fowlpox virus (FPV) transfer vector pUTA2. The recombinant plasmid and wild-type FPV 282E4 strain were then co-transfected into CEF cells and homologous recombination occurred. A recombinant virus expressing HIV-1 protein MEGp24 was screened by genome PCR and Western blot assay. Large scale preparation and purification of the recombinant fowlpox virus (rFPV) were then carried out. BALB/c mice were immunized intramuscularly with the rFPV for three times on day 0, 14 and 42. Mice were executed and sampled one week after the third inoculation. Anti-HIV-1 antibody in serum and Th1 cytokines in the supernatant of cultured spleen cells were assayed by ELISA. The count of T lymphocyte subsets and the CTL activity of spleen lymphocytes were analyzed by flow cytometry and lactate dehydrogenase (LDH) release assay, respectively. The results showed that HIV-1 specific antibody in serum and increased T lymphocyte subsets (CD4(+) T, CD8(+) T) were detected in the immunization group. CTL target-killing activity and higher secretion of Th1 cytokines (IFN-gamma and IL-2) of spleen lymphocytes stimulated by H-2(d)-restricted CTL peptide were observed in immunized mice. We concluded that the rFPV may induce HIV-1 specific immunity especially cellular immunity in mice.

Evaluation of recombinant fowlpox virus expressing infectious bronchitis virus S1 gene and chicken interferon-γ gene for immune protection against heterologous strains

“…The mixture was incubated in the dark at 4 • C for 30 min and then washed twice, followed by addition of 1.0 mL of PBS. The stained cells were detected by flow cytometry to count the T lymphocytes [13,14].…”

Immune responses of mice to prime-boost vaccination with the recombinant DNA and Fowlpox virus both expressing HIV-2 Gag-gp105

“…DNA vaccines have a significant advantage in the ability to elicit the CTL response (Herd et al, 2007;Molder et al, 2009). Moreover, the recombinant modified vaccinia virus Ankara (Ondondo et al, 2006;Wyatt et al, 2008) and recombinant FPV (Zhang et al, 2007;Emery et al, 2007) were reported to express HIV-1 proteins that elicited CTLmediated responses after the immunization.…”

Poxviral promoters for improving the immunogenicity of MVA delivered vaccines

“…16 Another example is the late ATI promoter (driving the A25L ORF), found in both VACV and cowpox virus. 29 The cowpox ATI promoter was utilised successfully in recombinant FPV 30 and MVA. 31 However, if the level of expression is very critical, a comparison should be performed to assess the activity of cross-stain exogenous promoters.…”